LOCUS       sequence-493104-        5686 bp    DNA     circular SYN 18-NOV-2025
DEFINITION  Final circuit with -Ara: OFF, +Ara: ON behaviour.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 24 to 58)
  AUTHORS   Nevot G, Guell M, Santos-Moreno J
  TITLE     Critical Analysis of Preprints and Inquiry-Based Lessons Improve the
            Synthetic Biology Learning Experience.
  JOURNAL   ACS Synth Biol. 2025 Aug 15;14(8):2878-2884. doi: 
            10.1021/acssynbio.5c00014.
  PUBMED    40814254
REFERENCE   2  (bases 1 to 5686)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Nov 18, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: "10.1021/acssynbio";
            journalName: "ACS Synth Biol"; date: "2025-08-15- 15"; volume: "14";
            issue: "8"; pages: "2878-2884"
COMMENT     Sequence Label: pEND-ZY
FEATURES             Location/Qualifiers
     source          1..5686
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(24..58)
                     /label=﻿BBa_J23100
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 1.0
                     "
                     /citation=[1]
     protein_bind    complement(427..446)
                     /label=Csy4 Site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease (Cermak 
                     et al. 2017)"
     protein_bind    complement(553..572)
                     /label=Csy4 Site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease (Cermak 
                     et al. 2017)"
     promoter        complement(633..917)
                     /gene="araBAD"
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the 
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     primer_bind     complement(691..710)
                     /label=pBAD Forward
                     /note="For vectors with E. coli araBAD promoter, forward 
                     primer"
     CDS             944..1822
                     /codon_start=1
                     /gene="araC"
                     /product="L-arabinose regulatory protein"
                     /label=araC
                     /translation="MAEAQNDPLLPGYSFNAHLVAGLTPIEANGYLDFFIDRPLGMKGY
                     ILNLTIRGQGVVKNQGREFVCRPGDILLFPPGEIHHYGRHPEAREWYHQWVYFRPRAYW
                     HEWLNWPSIFANTGFFRPDEAHQPHFSDLFGQIINAGQGEGRYSELLAINLLEQLLLRR
                     MEAINESLHPPMDNRVREACQYISDHLADSNFDIASVAQHVCLSPSRLSHLFRQQLGIS
                     VLSWREDQRISQAKLLLSTTRMPIATVGRNVGFDDQLYFSRVFKKCTGASPSEFRAGCE
                     EKVNDVAVKLS"
     rep_origin      1973..2608
                     /direction=RIGHT
                     /label=ColA ori
                     /note="Plasmids containing the ColA origin of replication 
                     can be propagated in E. coli cells that contain additional 
                     plasmids with compatible origins."
     primer_bind     2444..2463
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2626..2717
                     /gene="bla"
                     /label=AmpR promoter
     CDS             2718..3533
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             2718..3533
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(2791..2810)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      complement(3721..3748)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     protein_bind    complement(4068..4087)
                     /label=Csy4 Site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease (Cermak 
                     et al. 2017)"
     protein_bind    complement(4194..4213)
                     /label=Csy4 Site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease (Cermak 
                     et al. 2017)"
     promoter        complement(4357..4391)
                     /label=BBa_J23105
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.24"
     promoter        complement(4357..4391)
                     /label=BBa_J23107
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.36"
     promoter        complement(4357..4391)
                     /label=BBa_J23101
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.7"
     CDS             complement(4799..5512)
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     RBS             5520..5542
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    complement(5547..5566)
                     /label=Csy4 Site
                     /bound_moiety="Pseudomonas aeruginosa Csy4 (Cas6f)"
                     /note="cleavage site for the Csy4 endoribonuclease (Cermak 
                     et al. 2017)"
ORIGIN
        1 ggctatctcg ggcgtcccag cgagctagca ctgtacctag gactgagcta gccgtcaacc
       61 ggaccgcagg ctgaagctta ttttctacag tctcagtttc agggatttcg ttcactttac
      121 cttcttactc aataacaacc tatctactac caggcaccag ttctcgcaac agtgtatcaa
      181 ttaccaactt actgaaatca cagaactggg tgggactcgg cagagaaaac tcaatcggag
      241 actactttac tatctattat tcgttacagg tcagaagaat tgagaagaga aaagaaaacc
      301 gccgatcctg tccaccgcat tactgcaagg tagtggacaa gaccggcggt cttaagtttt
      361 ttggctgaac catggtaacg gagaagccct atcaactgtt tattgctcga gcgggcgtcc
      421 cagcgactgc ctatacggca gtgaacgcac cgactcggtg ccactttttc aagttgataa
      481 cggactagcc ttatttcaac ttgctatgct gtttccagca tagctctgaa acagtactta
      541 gtacacactg atctgcctat acggcagtga acccggaccg caggctggag ctcggacggt
      601 aaagtttcca gaatctatcc tatgaattcg gtatggagaa acagtagaga gttgcgataa
      661 aaagcgtcag gtaggaaccg ctaatcttat ggataaaaat gctatggcat agcaaagtgt
      721 gacgccgtgc aaataatcaa tgtggacttt tctgccgtga ttatagacac ttttgttacg
      781 cgtttttgtc atggctttgg tcccgctttg ttacagaatg cttttaataa gcggggttac
      841 cggtttggtt agcgagaaga gccagtaaaa gacgcagtga cggcaatgtc tgatgcaata
      901 tggacaattg gtttcttctc tgaatggcgg gagtatgaaa agtatggctg aagcgcaaaa
      961 tgatcccctg ctgccgggat actcgtttaa tgcccatctg gtggcgggtt taacgccgat
     1021 tgaggccaac ggttatctcg atttttttat cgaccgaccg ctgggaatga aaggttatat
     1081 tctcaatctc accattcgcg gtcagggggt ggtgaaaaat cagggacgag aatttgtttg
     1141 ccgaccgggt gatattttgc tgttcccgcc aggagagatt catcactacg gtcgtcatcc
     1201 ggaggctcgc gaatggtatc accagtgggt ttactttcgt ccgcgcgcct actggcatga
     1261 atggcttaac tggccgtcaa tatttgccaa tacggggttc tttcgcccgg atgaagcgca
     1321 ccagccgcat ttcagcgacc tgtttgggca aatcattaac gccgggcaag gggaagggcg
     1381 ctattcggag ctgctggcga taaatctgct tgagcaattg ttactgcggc gcatggaagc
     1441 gattaacgag tcgctccatc caccgatgga taatcgggta cgcgaggctt gtcagtacat
     1501 cagcgatcac ctggcagaca gcaattttga tatcgccagc gtcgcacagc atgtttgctt
     1561 gtcgccgtcg cgtctgtcac atcttttccg ccagcagtta gggattagcg tcttaagctg
     1621 gcgcgaggac caacgtatca gccaggcgaa gctgcttttg agcaccaccc ggatgcctat
     1681 cgccaccgtc ggtcgcaatg ttggttttga cgatcaactc tatttctcgc gggtatttaa
     1741 aaaatgcacc ggggccagcc cgagcgagtt ccgtgccggt tgtgaagaaa aagtgaatga
     1801 tgtagccgtc aagttgtcat aattggtaac gaatcagacc ctaggagcaa cccagtcagc
     1861 tccttccggt gggcgcgggg catgactaac atgagaatta caacttatat cgtatggggc
     1921 tgacttcagg tgctacattt gaagagataa attgcactga aatctagagt gatggtgtcg
     1981 ggaatccgta aaggatcttc ttgagatcct tttacgatcg tcgtaatctc ctgctctgta
     2041 aacgaaaaaa ccgcctgggg aggcggtttg atcgaaggtt aagtcagttg gggaactgct
     2101 taacctggta actggcttta gtggagcgca gataccaaat actgtccttt cagtgtagcc
     2161 tctgttaggc caccacttca agactctcga tatctaaatc cactaattct cagttaccaa
     2221 tggctgctgc cagtggcgtt ttgtcgtgtc tttccgggtt ggactcaaga tgatagttac
     2281 cggataaggc gcagcagtcg ggctgaacgg ggggttcttg cacacagccc agcttggagc
     2341 gaactgtcta cacggaacgg gacgtggtga tttgggtaaa gcctccacca caacacggac
     2401 gccgcaggac gggaacagga gagcgcaaga gggagccatc agggggaaac gcctggtatc
     2461 tttatagtcc tgtcgggttt cgccaccact gatttgagcg tcagatttcg tgatgttcgt
     2521 caggggggcg gagcctatgg aaaaacggct tcgctccggc cttattgtct ctctgctaag
     2581 tatcctcctg gcatcttcta ggacgtttct gcgctagcat gcctatttgt ttatttttct
     2641 aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat
     2701 attgaaaaag gaagagtatg agccatattc aacgggaaac gtcttgctct aggccgcgat
     2761 taaattccaa catggatgct gatttatatg ggtataaatg ggctcgcgat aatgtcgggc
     2821 aatcaggtgc gacaatctat cgattgtatg ggaagcccga tgcgccagag ttgtttctga
     2881 aacatggcaa aggtagcgtt gccaatgatg ttacagatga gatggtcaga ctaaactggc
     2941 tgacggaatt tatgcctctt ccgaccatca agcattttat ccgtactcct gatgatgcat
     3001 ggttactcac cactgcgatc cccgggaaaa cagcattcca ggtattagaa gaatatcctg
     3061 attcaggtga aaatattgtt gatgcgctgg cagtgttcct gcgccggttg cattcgattc
     3121 ctgtttgtaa ttgtcctttt aacagcgacc gcgtatttcg tctcgctcag gcgcaatcac
     3181 gaatgaataa cggtttggtt gatgcgagtg attttgatga cgagcgtaat ggctggcctg
     3241 ttgaacaagt ctggaaagaa atgcataaac ttttgccatt ctcaccggat tcagtcgtca
     3301 ctcatggtga tttctcactt gataacctta tttttgacga ggggaaatta ataggttgta
     3361 ttgatgttgg acgagtcgga atcgcagacc gataccagga tcttgccatc ctatggaact
     3421 gcctcggtga gttttctcct tcattacaga aacggctttt tcaaaaatat ggtattgata
     3481 atcctgatat gaataaattg cagtttcatt tgatgctcga tgagtttttc taagaattaa
     3541 ttcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     3601 acatttcccc gaaaagtgcc acttgcggag acccggtcgt cagcttgtcg tcggttcagg
     3661 gcagggtcgt taaatagccg cttatgtcta ttgctggttt accggtagat ctgaaacgca
     3721 aaaaggccat ccgtcaggat ggccttctgc ttagtttgat gcctggcagt ttatggcggg
     3781 cgtcctgccc gccaccctcc gggccgttgc ttcacaacgt tcaaatccgc tcccggcgga
     3841 tttgtcctac tcaggagagc gttcaccgac aaaggcgcgc cgctgaggac ctgtgtaaca
     3901 ggcgcctagt agagaaagta tttatcctca acagtgactc gtcgttcgct acctttggtc
     3961 gaaaaaaaaa gcccgcactg tcaggtgcgg gcttttttct gtgtttccgc ggccgcatct
     4021 tcagtttcca acgacgagtg taataggacg tccgggcgtc ccagcgactg cctatacggc
     4081 agtgaacgca ccgactcggt gccacttttt caagttgata acggactagc cttatttcaa
     4141 cttgctatgc tgtttccagc atagctctga aaccactaga gtctagcttg agactgccta
     4201 tacggcagtg aacccggacc gcaggctgtt caacagtatt gggtctgata agcactaccg
     4261 ggcgtcccag cgaatcagtg tgtactaagt acttggccgg accgcaggct gaatcgtggt
     4321 ttcaatcaga gcaactgttc acgggcgtcc cagcgagcta gcataatacc taggactgag
     4381 ctagccgtaa accggaccgc aggctggtcg actgagtttt caacgattac caatacttct
     4441 gactggattt cctgagcggt tctcttcacg caactacgag cacctactgt gtcttattac
     4501 aatctatctg ttatcgctac cctgtcccac tgaaaagtct acggaactgc agagtattat
     4561 tgatttacct gcgagaacga acaacactac cagacctatc ctttatctat taggaccaaa
     4621 acgaaaaaag gccccccttt cgggaggcct cttttctgga atttggtacc gagcctgcag
     4681 gagacagtgc tcaatcgtgt agaaatctct tggatcccgg gcgtcccagc gattattagt
     4741 cttcaatcca atccaggatg ctgtggattg tgatggcgtc agtattacgg cgggacattt
     4801 tatacagttc gtccatgccg tgggtgatac ccgcagccgt cacaaattcc agcaggacca
     4861 tgtggtcacg tttttcgttc ggatcttttg acagcacgga ctgggtactc aggtagtgat
     4921 tgtccggcag caggaccggg ccatcaccaa tcggcgtgtt ttgctgatag tggtcggcca
     4981 gctgaacgct accatcttcc acattatggc ggattttgaa attggcttta atgccgtttt
     5041 tctgtttatc cgcggtgatg tacacgttgt gggaattgaa gttatattcc agtttatgac
     5101 ccagaatatt gccgtcttct ttgaaatcga tacctttcag ttcgatacgg ttaaccaggg
     5161 tatcgccttc gaatttcact tccgcgcggg ttttatacgt accatcgtct ttaaagctaa
     5221 tcgtacgttc ctgcacatag ccttccggca tggccgattt gaaaaagtcg tgctgtttca
     5281 tgtgatccgg gtaacgcgca aaacattgaa cgccataggt cagcgtggtc accagcgtcg
     5341 gccacgggac cggcagttta cccgtggtgc agataaattt cagggtcagt ttgccgttcg
     5401 tcgcatcgcc ttcaccttcg ccgcgaacac tgaatttatg accattaaca tcgccgtcca
     5461 gttccaccag aatcggaacc acaccggtaa acagttcttc gcctttacgc atatgtatat
     5521 ctccttctta aagttaaaca aaattactgc ctatacggca gtgaacccgg accgcaggct
     5581 gtaagggaga ctttatcctc acagagttct acgggcgtcc cagcgatctc aagctagact
     5641 ctagtgtggc cggaccgcag gctgaaataa gtgtcacctg tgtaac
//
